The long-term goal of the proposed research is to determine the size and character of the molecular contractile stroke. This goal is approached in the short term by measurement of distinct steps of shortening in the single sarcomere of the single myofibril, a technique that has come into fruition in this laboratory. Because of high cooperativity among parallel thin filaments, translation in the myofibrillar sarcomere occurs in steps, apparently of the same size range as those found in optical trap experiments on single filaments. Thus, the stepping pattern in single myofibrils may reflect the step produced by the contractile stroke. Shortening steps in the single myofibril will be measured at the level of the single sarcomere or single half-sarcomere, at constant load. The steps are detected using one of two techniques: a photodiode array or a scanning galvanometer mirror plus photomultiplier. Step size will be determined under a variety of physiological conditions to determine how size is affected by these conditions. From the results we will learn much about the nature of contractile dynamics in the single sarcomere, and possibly also about the size and character of the molecular stroke.